In addition to their known uses in diagnostics, antibodies have been shown to be useful as therapeutic agents. For example, immunotherapy, or the use of antibodies for therapeutic purposes has been used in recent years to treat cancer. Passive immunotherapy involves the use of monoclonal antibodies in cancer treatments. See for example, Cancer: Principles and Practice of Oncology, 6th Edition (2001) Chapt. 20 pp. 495-508. These antibodies can have inherent therapeutic biological activity both by direct inhibition of tumor cell growth or survival and by their ability to recruit the natural cell killing activity of the body's immune system. These agents can be administered alone or in conjunction with radiation or chemotherapeutic agents. Rituximab and Trastuzumab, approved for treatment of non-Hodgkin's lymphoma and breast cancer, respectively, are two examples of such therapeutics. Alternatively, antibodies can be used to make antibody conjugates where the antibody is linked to a toxic agent and directs that agent to the tumor by specifically binding to the tumor. Gemtuzumab ozogamicin is an example of an approved antibody conjugate used for the treatment of leukemia. Monoclonal antibodies that bind to cancer cells and have potential uses for diagnosis and therapy have been disclosed in publications. See, for example, the following patent applications which disclose, inter alia, some molecular weights of target proteins: U.S. Pat. No. 6,054,561 (200 kD c-erbB-2 (Her2), and other unknown antigens 40-200 KD in size) and U.S. Pat. No. 5,656,444 (50 kD and 55 kD oncofetal protein). Example of antibodies in clinical trials and approved for treatment of solid tumors include: Trastuzumab (antigen: 180 kD, HER2/neu), Edrecolomab (antigen: 40-50 kD, Ep-CAM), Anti-human milk fat globules (HMFG1) (antigen >200 kD, HMW Mucin), Cetuximab (antigens: 150 kD and 170 kD, EGF receptor), Alemtuzumab (antigen: 21-28 kD, CD52), and Rituximab (antigen: 35 kD, CD20).
The antigen targets of trastuzumab (Her-2 receptor), which is used to treat breast cancer, and cetuximab (EGF receptor), which is in clinical trials for the treatment of several cancers, are present at some detectable level on a large number of normal human adult tissues including skin, colon, lung, ovary, liver, and pancreas. The margin of safety in using these therapeutics is possibly provided by the difference in the level of expression or in access of or activity of the antibody at these sites.
In addition to cancer targets, antibody therapeutics have also been shown to be effective against chronic inflammation and other immune disorders. An example of an antibody therapeutic approved for treatment of immune disorders is Infliximab (antigen: TNFα).
Another type of immunotherapy is active immunotherapy, or vaccination, with an antigen present on a specific cancer(s) or a DNA construct that directs the expression of the antigen, which then evokes the immune response in the individual, i.e., to induce the individual to actively produce antibodies against their own cancer. Active immunization has not been used as often as passive immunotherapy or immunotoxins.
Several models of disease (including cancer) progression have been suggested. Theories range from causation by a single infective/transforming event to the evolution of an increasingly “disease-like” or ‘cancer-like’ tissue type leading ultimately to one with fully pathogenic or malignant capability. Some argue that with cancer, for example, a single mutational event is sufficient to cause malignancy, while others argue that subsequent alterations are also necessary. Some others have suggested that increasing mutational load and tumor grade are necessary for both initiation as well as progression of neoplasia via a continuum of mutation-selection events at the cellular level. Some cancer targets are found only in tumor tissues, while others are present in normal tissues and are up regulated and/or over-expressed in tumor tissues. In such situations, some researchers have suggested that the over-expression is linked to the acquisition of malignancy, while others suggest that the over-expression is merely a marker of a trend along a path to an increasing disease state.
An ideal diagnostic and/or therapeutic antibody would be specific for an antigen present on a large number of cancers, but absent or present only at low levels on any normal tissue. The discovery, characterization, and isolation of a novel antigen that is specifically associated with cancer(s) would be useful in many ways. First, the antigen could be used to make monoclonal antibodies against the antigen. An antibody would ideally have biological activity against cancer cells and be able to recruit the immune system's response to foreign antigens. An antibody could be administered as a therapeutic alone or in combination with current treatments or used to prepare immunoconjugates linked to toxic agents. An antibody with the same specificity but with low or no biological activity when administered alone could also be useful in that an antibody could be used to prepare an immunoconjugate with a radio-isotope, a toxin, or a chemotherapeutic agent or liposome containing a chemotherapeutic agent, with the conjugated form being biologically active by virtue of the antibody directing the toxin to the antigen-containing cells.
One aspect desirable for an ideal diagnostic and therapeutic antibody is the discovery and characterization of an antigen that is associated with a variety of cancers. There are few antigens that are expressed on a number of types of cancer (e.g., “pan-cancer” antigen) that have limited expression on non-cancerous cells. The isolation and purification of such an antigen would be useful for making antibodies (e.g., diagnostic or therapeutic) targeting the antigen. An antibody binding to the “pan-cancer” antigen could be able to target a variety of cancers found in different tissues in contrast to an antibody against an antigen associated with only one specific type of cancer. The antigen would also be useful for drug discovery (e.g., small molecules) and for further characterization of cellular regulation, growth, and differentiation.
The ADAMs (ADisintegrin and Metalloprotease Domain) are a family of proteins thought to be important in a wide range of biological processes such as cell adhesion, cell fusion and proteolysis. In general, they are transmembrane proteins having homology to snake venom proteins that have the ability to degrade basement membrane component and bind to integrins, thus disrupting cell-matrix interactions. Because of the similarity in function to snake venom proteins, ADAMs are also known as MDC proteins (Metalloprotease-like, Disintegrin, Cysteine-rich).
MDC9 or ADAM-9 (Meltrin γ) is a member of the ADAM/MDC protein family. The protein sequence has been described in U.S. Patent Application Nos. 20040091473 and 20020042368. ADAM-9 expression has been found in rat kidney and also been shown to be up-regulated in hepatocellular carcinomas using protein microarrays, however the significance of this finding to cancer progression is unclear. ADAM-9 has also been shown to be involved in the shedding of heparin-binding epidermal growth factor (HB-EGF), in the processing of the amyloid precursor protein, and as a ligand for several integrins. Although it is highly conserved between mouse, human and Xenopus and it is widely expressed in different tissues, a knock-out mouse lacking ADAM-9 shows no evident pathological abnormalities. Polyclonal and monoclonal antibodies against ADAM-9 have been used in the identification of novel proteins associated with hepatocellular carcinoma in protein arrays (Tannapfel et al., J of Pathology 2003; 201: 238-49) and also in atherosclerosis models using venous vascular smooth muscle cells (VSMCs) to assay for ADAM-9 expression (Al-Fakhri et al. J Cellular Biochem (2003)89: 808-823), however the significance of ADAM-9 as a marker is unclear. This prior art does not teach or suggest ADAM-9 antibodies as therapeutic agents, or that the expression of ADAM-9 plays a causative effect in cancer.
What is needed are novel targets on the surface of diseased and/or cancer cells that may be used to diagnose and treat such diseases and/or cancers with antibodies and other agents which specifically recognize the cell surface targets. There exists a further need, based on the discoveries disclosed herein, for novel antibodies and other agents which specifically recognize targets on the surface of cells that can modulate, either by reducing or enhancing, the disease-promoting activities of ADAM-9. It is an object of this invention to identify antagonists of human ADAM-9 that are capable of inhibiting its disease-associated activities. It is another object to provide novel compounds for use in the assay of ADAM-9, and for use as immunogens or for selecting anti-human ADAM-9 antibodies.
As will be described in more detail below, the present inventors have made discoveries concerning the known polypeptide, ADAM-9, which we refer to herein as KID24, identified as the antigen target of the novel antagonists, modulators and antibodies provided herein.
All references, publications and patent applications disclosed herein are hereby incorporated by reference in their entirety.